The paper is how to transform any cell into a stem cell. In the paper the authors took an “I’m so smart” approach to the whole problem. These smart guess approaches rarely work, but it’s always worth trying. This paper demonstrates why.

So let’s get back to the problem. Reprogramming cells so that they can become totipotent. How to proceed? From the paper:

We selected 24 genes as candidates for factors that induce pluripotency in somatic cells, based on our hypothesis that such factors also play pivotal roles in the maintenance of [embryonic stem] cell identity.

So here is a list of 24 genes that when activated COULD promote the reprogramming of cells so that they can acquire stem cell like properties. Then what? The researchers inserted the genes into vectors and then threw the mix of all 24 DNA constructs onto fibroblasts (mouse embryonic fibroblasts) that had an drug resistance marker controlled by a stem cell specific promoter. The idea is that if the MEFs became stemcells, they will turn on stemcell specific promoters and thus acquire resistance to the drug G418. All you have to do to find these “stemcells” is grow the cells in the presence of the drug and see whether any cells survive.

Sure enough they could find such cells. Wow! And not only that but these resistant cells formed colonies and according to the authors each colony

exhibited morphology similar to ES cells, including a round shape, large nucleoli, and scant cytoplasm.

Wow!!

The next thing to do is cut down the number of genes, after all 24 is quite a huge number.

And so they did … to four genes, each of which encodes a transcription factor.

So what are the identities of these genes?

Oct3/4, Klf4, Sox2 and c-Myc.

That’s it.

The authors wanted to then analyze cells that had been converted by the introduction of these four factors, now called iPS cells for induced pluripotent stem cells. They looked at overall gene expression in iPS vs embryonic stem cells, the activation of key stemcell markers in iPS cells and the ability of these cells to differentiate into other cell types. As far as the authors could tell, these cells looked like stemcells. From the paper:

Comments

The main issues are going to be how far can these cells be expanded ?(they went as far as 30 passages which is pretty good). Also this must be repeated with human cells, and ideally, get away from retroviral transduction which has led to problems with oncogenesis in the past. Maybe an integrase system?

There was a lot of concern about the validity of this paper, not because the experimental design was faulty, but because the results were so incredible. In less than a week all these concerns will vanish.

This is just the tip of the iceberg. Tthink of it – you get 4 transcription factors turned on and presto, you’ve made stem cells. Potentially if you needed stem cells you could just take a swab of cells from your mouth, microinject the 4 proteins into them and *poof* stemcells. Now of course the process of conversion is very inefficient, but at least we have some sort of handle on how to reprogram cells. My guess is that the more we know, the better we’ll get at it. At some point this will surely lead to certain therapies for many diseases – faster than anyone imagined.

This is a cool paper and it’s exciting to be able to finally identify the key transcription factors that can reprogram a cell.

But, not to put a damper on the significance of the work, it’s hardly a surprise. At least not in the sense of revealing any fundamental new principles of biology.

Why is this relevant? Because over on Sandwalk I’m covering the top 25 science questions from Science magazine in June 2005. One of them was How Can a Skin Cell Become a Nerve Cell?. The science journalists who made up the questions thought that this problem was one of the fundamental mysteries of biology. I think they were wrong … very wrong.

Why is there such a huge disconnect between what science writers for Science think is a major problem and what actual scientists think? Are there any actual scientists who think that this paper revealed something we didn’t know already; namely that transcription factors control development? You don’t really think it will win a Nobel Prize for the authors, do you?

Larry, I think it was more impressive because with the four factors they were able to reverse all the epigenetic programming which had seemed somewhat irreversible in an adult cell. While we new from oocyte programming something similar could be done, narrowing it down from a field of 20 candidates to 4 factors really was pretty cool. And that the factors can eliminate epigenetic programming is pretty extraordinary.

You must be joking … I guess using your logic we can write off any finding concerning gene programs with the quip “it’s no surprise that transcription factors regulate genetic programs.” The big finding is that it is easy to dedifferentiate a cell, something totally unexpected. During development in vertebrates and other organisms, the germ cell lines are segregated from the rest of the embryo. It was thought by some that this was due to the fact that it is just hard to make germ cells from somatic cells. This work by Takahashi and Yamanada shows that this is not the case. This work, just like Hartwell and Nurse’s work on the cell cycle, is a major breakthrough in that we now have a way to examine how this genetic program works. Also if you think about it, cloning just became a whole lot easier.

The big finding is that it is easy to dedifferentiate a cell, something totally unexpected. During development in vertebrates and other organisms, the germ cell lines are segregated from the rest of the embryo early on. It was thought by some that this was due to the fact that it is just hard to make germ cells from somatic cells.

Why do you say this was “totally unexpected?” It sure wasn’t for me. I learned about Gurdon’s experiments with frog cells back when I was an undergraduate in the 1960’s. We’ve known for over 40 years that it’s possible to reprogram differentiated animal cells.

Now we’re learning more about the molecular basis of that process and that’s cool. But it’s hardly a surprise that it has something to do with transcription factors.

Gurdon’s work was with nuclear cell transfers – I’m not sure how much he or anyone else progressed from there. And nuclear transfer has also been employed to produce ES cells and clones in other systems. But this technique is hard to perform with mammalian oocytes and the technique did not provide insights as to how cells dedifferentiated. For example did we learn how many oocyte factors were involved in this reprograming process? What was the nature of those factors? All a big mystery.

However in this paper that I describe, we find out that expressing four genes can lead to a total reprogramming of cells – now that is a tremendous advance. You must admit that. Sure a genetic program was bound to be involved, but no one would have though that the introduction of four factors could do it. You may think that this is not a “big advance” but I’m afraid that you are in the minority. With all the effort and money being poured into stemcell research and all the potential that is yet to be tapped from the findings in this field, it is almost assured that this paper will be the basis for a Nobel … much more so than the human genome project.

An undeniable advance. Reading your review of the paper akes me recollect how fascinated I was when I read the paper. Its funny, so I work in a lab in physicists so when the paper came out and I expressed my enthusiasm about it, everyone just gave me empty stares!

Reminds me of the “minimal requirements for cancer” several years back by Weinberg’s group that showed that only Ras, large T, and hTert were sufficient for transformation of normal cells. So simple. Too simple.

Well, I’m sure like everything else in biology, in “real world applications” it’ll turn out to be more complicated. But I am nontheless excited to read this and the forthcoming papers on the subject.
It seems like the most kick-ass experiments are like the ones in this paper: a lot of work, but you wonder how is it that no one has done this before?

With all the interest I see here in stem cells I think it is time to delve into the prospects of designing do it yourself kits so those of us who are amature scientists might be able to do the same extraction and processing.
Equipment seems to be reasonable in price and redily available.

Let’s do it. Joe

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